Homofermented products

ABSTRACT

The present invention pertains to the field of liquid feed, and relates to fermented feed products, methods for their preparation, as well as uses thereof. Furthermore, the invention further relates to animal liquid feed and methods of preparing liquid feed using lactic acid bacteria. In particular, a method of preparing a fermented mixed feed is provided, comprising the steps of: (a) providing a liquid fermented product; (b) providing a feed product to be fermented; (c) combining the products from step (a) and (b), and fermenting the feed product of step (b) using the liquid fermented product of step (a) as inoculum.

TECHNICAL FIELD OF THE INVENTION

The present invention pertains to the field of liquid feed. Moreparticularly, the present relates to providing fermented feed products,methods for their preparation as well as uses thereof.

BACKGROUND OF THE INVENTION

In the farming industry, feed is often delivered to the animals byliquid feeding systems. This causes several problems. Potential harmfulbacteria and organisms are natural inhabitants of soil and vegetationand are accordingly found on feed components and everywhere in theanimal's surroundings. The bacteria and other organisms present willferment, unless prevented e.g. by sterilisation. The fermentation mayresult in outgrow of pathogenic bacteria e.g. Vibrio spp., Campylobacterspp., Salmonella spp., E. coli, and Stapholococcus aureus. Furthermore,there might be a high content of various types of yeast and moulds. Thisuncontrolled growth in the liquid feed may result in illness,malnutrition, diarrhea, or even death of the animals. Furthermore,animals infected with Campylobacter spp. or Salmonella spp. may transferthe infection to humans, and therefore it is desirable to avoid suchinfections in animals.

Production of a fermented mixed feed in an environment with highmicrobial pressure, including pathogenic microorganisms is verychallenging, and many attempts have failed so far. Generally,conventional inoculation of feed with starter cultures requirescultivation and propagation steps, including a sterile handling ofbacteria. This is highly challenging for a farmer, who is not trained ingeneral microbiological skills, and who is surrounded by an environmentof high microbial pressure, such as found in conventional feedingkitchens in a pig farm.

The alternative, a “continuous” fermentation, where a portion of thefermented feed is used as inoculum for a new fermentation batch, suffersfrom gradual increase of undesired microorganisms, such as acid tolerantyeasts. These are not only known to have a detrimental effect on thefermented feed, but they can be very difficult to get rid off,especially in the feeding pipes.

It has proven very difficult, if not impossible, to avoid theabove-mentioned problems if liquid feed is to be used as feed.Accordingly, the use of dry feed is often the alternative choice.

Until recently, antibiotic-based growth promoters were provided to e.g.pigs on a routine basis, in order to prevent e.g. diarrhea, and therebyto increase weight gain (hence the term “growth promoter”). The use ofantibiotic-based growth promoters by farmers has been banned in severalcountries, including EU. This is in part caused by the fear ofgenerating multi-resistant pathogenic microorganisms. However, use ofantibiotics has not stopped, and veterinarians prescribe antibiotics fortreatment of large numbers of pigs every day.

By the present invention, the above-mentioned problems have been solved.Furthermore, the invention provides a non-antibiotic growth promoter.

SUMMARY OF THE INVENTION

In one aspect, the present invention relates to a method of preparing afermented mixed feed, comprising the steps of: (a) providing a liquidfermented product; (b) providing a feed product to be fermented; (c)combining the products from step (a) and (b), and fermenting the feedproduct of step (b) using the liquid fermented product of step (a) asinoculum.

Another aspect of the invention concerns a fermented mixed feed,provided by a method comprising the steps of: (a) providing a liquidfermented product; (b) providing a feed product to be fermented; (c)combining the products from step (a) and (b), and fermenting the feedproduct of step (b) using the liquid fermented product of step (a) asinoculum.

A further aspect of the invention pertains to the use of a fermentedmixed feed for feeding animals, said fermented mixed feed being providedby a method comprising the steps of: (a) providing a liquid fermentedproduct; (b) providing a feed product to be fermented; (c) combining theproducts from step (a) and (b), and fermenting the feed product of step(b) using the liquid fermented product of step (a) as inoculum.

A further aspect of the invention relates to fermented products whichcan be obtained by fermenting a fluid product with lactic acid-producingbacteria capable of producing sufficient lactic acid to decrease the pHto at least 4.2 within 24 hours.

In a further aspect, the invention relates to silage, its preparationand use.

In a further aspect, the present invention relates to uses of thefermented products according to the invention either directly as liquidfeed or for preparing liquid feed.

In a further aspect, the present invention relates to a method forpreparing a fermented product and its use as growth promoter. The methodcomprises: (a) providing a fluid product; (b) providing lacticacid-producing bacteria capable of producing sufficient lactic acid todecrease the pH to at least 4.2 within 24 hours; (c) bringing (a) and(b) together; and (d) allowing fermentation to take place under suitableconditions to obtain said fermented product.

In a further aspect, the present invention relates to a method ofpreparing a liquid feed comprising (a) providing a fermented productaccording the present invention; (b) providing a feed product to befermented; and (c) allowing fermentation to take place under suitableconditions.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows the homofermentative process as compared to theheterofermentative process.

FIG. 2 shows at which pH lactic acid-producing bacteria is active and atwhich pH some of the unwanted bacteria is able to grow.

FIG. 3 is a schematic presentation of a well-controlled naturalfermentation process.

FIG. 4 is a schematic presentation of a poorly controlled naturalfermentation process.

FIG. 5 shows the effect on the cadaverine production (rotting process),depending on whether the fermentation process is controlled or not.

FIG. 6 is a flow diagram, showing the steps of providing animal feedcomprising a fermented product or by-product from an industrial processaccording to an embodiment of the invention.

FIG. 7 is a flow diagram, showing the steps of providing a fermented,liquid animal feed according to an embodiment of the invention.

FIG. 8 shows illustrates main components and steps according to anembodiment of the invention.

FIG. 9 shows the production of fermented feed according to analternative embodiment of the invention.

FIG. 10 is a schematic representation of a setup with fermentation andmixing tank according to an embodiment of the invention.

FIG. 11 is a schematic representation of a setup with fermentation tankaccording to an embodiment of the invention.

FIG. 12 is a schematic representation of a setup with severalfermentation tanks according to an embodiment of the invention.

DETAILED DESCRIPTION OF THE INVENTION

Preparation of liquid feed involves a fermentation or ensiling process.In general, two types of fermentation processes can take place, namely“heterofermentation” by which the fermentation results in the formationof lactic acid, acetic acid, butyric acid, yeast and ethanol, and“homofermentation” by which the fermentation results in the formation oflactic acid.

Thus, it is an object of the present invention to control thefermentation process by ensuring that homofermentation is primarilytaking place and by ensuring that the growth of unwanted bacteria,yeast, moulds, fungi and lactic acid-producing bacteria are controlledor outcompeted.

It is well-known that different acids and yeasts have different tastes,some of which the animals may like and some which they dislike.

Thus, it is a further object of the present invention to ensure thecorrect components are present and that such components are present in asuitable ratio resulting in higher palatability.

The lactic acid bacteria according to the invention are capable ofproducing lactic acid and decreasing the pH during fermention to atleast 4.2 within 24 hours. In one embodiment, the lactic acid-productionshould take place within 15 hours.

In one embodiment, the pH is from within 3.5 to 4.2 such as e.g. 3.8.

In one aspect, the present invention relates to a fermented productwhich is obtainable by fermenting a fluid product with lacticacid-producing bacteria capable of producing sufficient lactic acid todecrease the pH to at least 4.2 within 24 hours. In one embodiment, thepH is between 4.2 and 3.5 such as 3.8. In another embodiment, thedesired pH should be reached within 15 hours or less.

In the present context, the terms “fermentation” and “ensiling” is usedinterchangeably and is intended to have the same meaning.

The terms “fermenter” or “fermentation tank” indicate a container, atank or the like, in which fermentation takes place. Commonly, one ormore mixing devices for mixing the content of the fermenter areprovided. However, fermentation or a part of fermentation can also takeplace outside of a fermenter, such as within a pipe, mixing tank,storage tank and the like.

The terms “tank” or “container” are interchangeable, and indicate a(storage) facility. They can be of silo-type, big-bags, or a storagefacility, such as a room or separate compartment.

The terms “fluid product” or “liquid product” are used interchangeablyand indicate a product with a moisture content of 20 vol-% or more, inparticular 25 vol-% or more. Within the present context, “vol-%” isintended to mean volume percent. Unless indicated differently, %indicates weight %.

In order to be suitable for fermentation according to the invention, aproduct will commonly have a moisture content of 20 vol-% or more, inparticular 25 vol-% or more. In one embodiment of the invention, asufficient water content (i.e. a water content of 20 vol-% or more) isprovided by mixing a product with an insufficient water content (i.e. awater content of below 20-vol %), with a product with a sufficient watercontent, in order to provide an appropriate water content of themixture. In a further embodiment, a combination of mixing products withdifferent water contents combined with addition of water or awater-based fluid provides a mixture with appropriate water content forfermentation.

In another embodiment of the invention, water is added to the feedproduct to be fermented, if the moisture content is too low forefficient fermentation. Optionally, the water can be treated, and cancomprise chemical compounds and chemical compositions, such as salts,minerals, vitamins, buffering substances, organic or inorganic acids andthe like. In yet another embodiment, the treated water improvesfermentation and/or weight gain in animals.

The term “product” according to the invention is to be understood in itsbroadest sense. Commonly, the products are food or feed related.“Product(s)” and “feed product(s)” may suitably be obtained from thedairy industry, the agricultural industry, the wine industry, the spiritindustry, or beer industry, or combinations thereof. Examples ofsuitable “products” and “feed products” comprise one or more of matureand/or immature plants and parts thereof, such cereals, e.g. wheat,barley, rye, rice, maize (cob maize silage (CCM) or ripe), triticale,oat; vegetables (e.g. potatoes, beans, peas, maize, soy; whey, curd,skim milk and the like.

“By-products” or “waste products” are terms used to describe mostlyundesired products deriving from industrial processes, which may beavailable at low costs, or for free. Commonly, they are not useddirectly as feed for animals, and long time storage may be an issue dueto decomposition and uncontrolled fermentation and spoilage. Examples ofsuch “by-products” and/or “waste products” comprise whey, spent grain(from brewing, wine or bio-ethanol industry), plant or parts thereof,potatoes, plants, yeast, bacteria, fungi and the like.

The terms “fermented product” or “fermented feed” indicate any productor feed that has been fermented or is in the process of being fermented.

“Liquid fermented product” indicates that the fermented product has awater/moisture content of more than 20%. In the context of the presentinvention, it is used as inoculum for fermentation. It can also be fedto animals directly, or in combination with another fermented ornon-fermented feed, composition or product.

“Silage” or “ensilage” are used interchangeably and refer to a fermentedplant product to be fed to animals, commonly ruminant animals likecattle and sheep. Silage is fermented and stored, a process calledensilage. Ususally, fermentation occurs via the natural microbial florapresent on the plant product to be fermented. Thereby, a variety ofdifferent fermentation products can occur, including acetic acid. Thefermentation process can take days, weeks or month, and the resultingfermented product, the silage or ensilage can be stored for many months.Silage is most often prepared from grass crops, including maize orsorghum. Silage is often made from the entire plant, usually excludingthe roots, and not just the grain. Generally, the plant is cut intopieces, often directly during harvesting. Silage can also be made frommany other field crops, and sometimes a mixture is used, such as oatsand peas. Haylage is a term used to describe ensiled forages, made up ofgrass, alfalfa and alfalfa/grass mixes and the like. It is used forexample to feed dairy. Instead of fermenting in a silo, baylage isanother form of silage. In this case, for example the plant or partsthereof is cut and baled while still fairly wet, and often too wet to bebaled and stored as hay. The dry matter can be around 60 to 70%. Thebales are wrapped tightly in plastic wrappers, wherein fermentationoccurs.

The term “fermented mixed feed” indicates a feed product that is in theprocess of or that has been fermented according to the invention by theuse of a liquid fermented product as inoculum. Additional components,e.g minerals, vitamins, amino acids, non fermented feed and the like,may be present as well.

The term “animal(s)” as used herein is intended to include mammals suchas pigs, piglets, cattle, and horses, poultry such as chickens, turkeys,hens, geese and ducks, and fish such as salmon and trout. Monogastricanimals, such as humans, pigs, horses, dogs, and cats, have a simplesingle chambered stomach. In contrast, ruminant animals or ruminantshave a multi-chambered complex stomach. Ruminants digests their food intwo steps, first by eating the raw material and regurgitating asemi-digested form known as cud, then eating (chewing) the cud, aprocess called ruminating. Ruminants include for example cattle, goats,sheep and deer.

The term “growth promoter” indicates a compound or product which isprovided to an animal in order to improve its growth or weight gain.This includes addition of antibiotics to the feed, either prescribed bya veterinarian or added prophylactically by a farmer. On the other hand,a fermented mixed feed provided by the invention is also a growthpromoter, as it has been shown to be able to increase weight gain ofanimals compared to a control.

In general, the fermentation takes at a suitable temperature for asuitable amount of time. The temperature should be so as to ensureactivity of the lactic acid-producing bacteria, i.e. it should not bebelow 10° C. or above 50° C. In one embodiment, the temperature may beabout 30° C. The fermentation should be continued at least until the pHhas reached a suitable level, i.e. 4.2 or less, e.g. 3.5 or 3.8. Thus,fermentation should be continued up to e.g. 24 hours, or 15 hour.Sometimes, fermentation may be completed within 10, 9, 8, 5, 6, 4, 3, or2 hours or 1 hour, or in certain cases within 30 minutes.

In one aspect, the present invention relates to a method of preparing afermented mixed feed, comprising the steps of: (a) providing a liquidfermented product; (b) providing a feed product to be fermented; (c)combining the products from step (a) and (b), and fermenting the feedproduct of step (b) using the liquid fermented product of step (a) asinoculum. In one embodiment of the invention, the fermented mixed feedhas a pH below 4.2. In another embodiment, the pH is between 4.2 and3.5. In a further embodiment, the pH of the fermented mixed feed isaround 3.8.

Lactic acid bacteria produce lactic acid during fermentation of afermentable carbon source, which results in acidification of theenvironment. Depending on the starter culture or starter cultures used,as well as on the availability of fermentable sugar(s), a fermentedmixed feed is provided according to the invention, said fermented feedpossesses a lactic acid concentration above 50 mM. In anotherembodiment, lactic acid concentration exceeds 100 mM. In a furtherembodiment, the lactic acid concentration lactic exceeds 150 mM or 200mM. In yet another embodiment, lactic acid concentrations of above 250mM or above 300 mM are provided in the fermented product.

The lactic acid concentration in the liquid fermented product,by-product or waste product used as inoculum for the fermentationaccording to the invention can be higher than the lactic acidconcentration in the fermented mixed feed. In another embodiment of theinvention, the lactic acid concentration in the fermented mixed feed ishigher than in the liquid fermented (by-)product. In a furtherembodiment, the lactic acid concentrations of inoculum and fermentedproduct are approximately the same.

Likewise, pH of incoculum and fermented (mixed) product can be the same,similar or different. In one embodiment of the invention, the pH of theliquid fermented product used as inoculum is below 4.2. In anotherembodiment, the pH is between 4.2 and 3.5. In a further embodiment, thepH is around 3.8.

According to the invention, a fermented mixed feed can be producedwithin 1 day, or within 12-24 h. In another embodiment, the fermentedproduct can be produced between 8-12 h, or 6-8 h. In a furtherembodiment, the fermentation is achieved between 4-6 h, or below 4 h.

In another aspect of the invention, fermentation can be slower, and cantake one or more days, several days, one week, several weeks, one month,or several months. Fermentation can be controlled by adding a fermentedliquid product or a mixed fermented feed as inoculum comprising activelactic acid bacteria. The proportion of inoculum to feed product to befermented can be in the range of 0.1-10 vol-%, or 0.5-5 vol-%, or1-2.5%, or around 1-2 vol-%. The fermentation process can take place ina closed silo. In another embodiment of the invention, fermentationoccurs in bales tightly wrapped in plastic wrappers. In one particularembodiment of the invention, ensilage is based on fermented CCM maize.The resulting ensilage can be used to feed monogastric animals. In afurther embodiment, CCM maize silage is suitable for feeding pigs. Inyet another embodiment, CCM maize silage can be used as growth promoterfor feeding monogastric animals, such as pigs. The CCM maize basedensilage can be low in acetic acid. In one embodiment of the invention,the concentration of acetic acid is 20 mM or lower, or 15 mM or lower,or 10 mM or lower, or 5 mM or lower. In a further embodiment of theinvention, lactic acid content is 50 mM or more, 100 mM or more, 150 mMor more, 200 mM or more, 250 mM or more, or above 300 mM.

Fermentations according to the invention can be performed at differenttemperature ranges. Commonly, fermentation is performed at a temperaturebetween 5° C. and 50° C., or between 15° C. 40° C. In another embodimentof the invention, fermentation temperature is between 18 and 30° C. In afurther embodiment of the invention, fermentation is conducted at oraround room temperature, such as 20-25° C., or 22-24° C. or around 23°C. In yet another embodiment of the invention, means for monitoring andcontrolling temperature are provided. In yet a further embodiment of theinvention, the temperature of the water added for providing anappropriate water content of the fermentation broth is essentiallycontrolling fermentation temperature. Fermentation temperature can beconstant, or it may vary.

Suitable fermenters according to the invention can be provided withmixing means, often provided as standard equipment from themanufacturer. Commonly, they will be operated at approximately 1-500rpm. In one embodiment of the invention, mixing speed is 10-300 rpm. Inanother embodiment of the invention, the mixing means is operatedbetween 20 and 100 rpm. In a further embodiment, the mixing deviceoperates at around 35 rpm.

According to the invention, a product to be fermented is combined with aliquid fermented product comprising (active) lactic acid bacteria. Thisinoculum can be present in a range of 0.1 to 99.9 vol-%, or 1 to 99vol-%, or 5 to 70 vol-%. In another embodiment of the invention, theinocolum is around 0.5-1, 1, 1-5, 5, 5-10, 10, 10-20, 20, 20-30, 30,30-40, 40, 40-50 or 50 vol-%. In a further embodiment, the liquidfermented product comprises 25 to 35 vol-%. In yet another embodiment,the inoculum comprises around 30 vol-%.

The fermentation process provided by the starter culture according tothe invention is essentially a homofermentative process (FIG. 1).“Essentially homofermentative” means, that the predominant bacterialflora driving the fermentation is homofermentative. In one embodiment,99% or more of the bacteria are homofermentative. In another embodimentof the invention, 95% or more of the bacteria are homofermentative. Inyet another embodiment, 90% or more of the bacteria arehomofermentative. “Essentially homofermentative” indicates also that themajor fermentation product is lactic acid, and the levels of acetic acidand ethanol are either below taste threshold, around taste threshold orslightly above taste threshold. Alternatively, “essentiallyhomofermentative” indicates a ratio of lactic acid to acetic acid orlactic acid to ethanol (mM/mM) of 10:1 or more, 20:1 or more, 50:1 ormore, or 100:1 or more. According to the invention, both fermentationsare essentially homofermentative, i.e. fermentation of the fermented(by-) product as well fermentation of the feed product (barley, wheat,soy etc.). In another embodiment of the invention, the fermentationprocess is a heterofermentative process (FIG. 1). In a furtherembodiment of the invention, the starter culture comprises homo- andheterofermentative lactic acid bacteria. In yet another embodiment ofthe invention, the starter culture comprises non-lactic acid bacteria.

According to the invention, fermentations comprise lactic acid-producingbacteria. Thus, in the broadest aspect, the present invention relates tolactic acid-producing bacteria. The lactic acid bacteria comprise a ladeof Gram positive, low-GC, acid tolerant, non-sporulating, non-respiringrod or cocci that are associated by their common metabolic andphysiological characteristics. These bacteria, usually found indecomposing plants and lactic products produce lactic acid as the majormetabolic endproduct of carbohydrate fermentation. This trait hashistorically linked lactic acid bacteria with food fermentations asacidification inhibits the growth of spoilage agents. Proteinaceousbacteriocins are produced by several lactic acid bacteria strains andprovide an additional hurdle for spoilage and pathogenic microorganisms.Furthermore, lactic acid and other metabolic products contribute to theorganoleptic and textural profile of a food item. The industrialimportance of the lactic acid bacteria is further evidenced by theirgenerally regarded as safe (GRAS) status, due to their ubiquitousappearance in food and their contribution to the healthy microflora ofhuman mucosal surfaces. The genera that comprise the lactic acidbacteria are Lactobacillus, Leuconostoc, Pediococcus, Lactococcus, andStreptococcus, Aerococcus, Carnobacterium, Enterococcus, Oenococcus,Teragenococcus, Vagococcus, and Weisella; these genera belong to theorder Lactobacillales.

In the present invention, the lactic acid-producing bacteria used forfermentation are mainly and non-exclusively lactic acid bacteria of thegenus Enterococcus, Lactobacillus, Pediococcus or Lactococcus, orcombinations thereof. A starter culture or inoculum according to theinvention can also comprise lactic acid bacteria selected from the groupconsisting of one or more Enterococcus spp., Lactobacillus spp.,Lactococcus spp., and Pediococcus spp. In another embodiment of theinvention, the lactic acid bacteria are selected from the groupconsisting of one or more one Enterococcus faecium, Lactobacillusrhamnosus, Lactobacillus plantarum, Pediococcus acidililactili, andPediococcus pentosaceus. In further embodiment, the lactic acidproducing bacteria are of the order Lactobacillales. The lacticacid-producing bacteria can also be selected from Lactobacillus spp.,Pediococcus spp., Enterococcus spp., and Lactococcus spp. or acombination thereof. In yet another embodiment, the lacticacid-producing bacteria comprise Pediococcus pentosaceus, Pendiococcusacidilactici and Lactobacillus plantarum, Lactobacillus rhamnosus, andEnterococcus faecium, or a combination thereof. In still anotherembodiment, the lactic acid bacteria comprise Enterococcus faeciumand/or Lactobacillus rhamnosus. In a further embodiment, the lactic acidbacteria comprise one or more of Enterococcus faecium MCIMB 30122,Lactobacillus rhamnosus NCIMB 30121, Pediococcus pentosaceus HTS (LMGP-22549), Pendiococcus acidilactici NCIMB 30086 and/or Lactobacillusplantarum LSI (NCIMB 30083).

According to one aspect of the invention, a product, by-product or wasteproduct is fermented. Such a product, by-product or waste product may beobtained from various industries, which may or may not be related tofeed and/or food production. Suitable industries may comprise foodindustry, feed industry, dairy industry, agricultural industry, alcoholproducing industry, beer industry, wine industry, distilled spiritindustry, bio-ethanol industry, meat processing industry, fishingindustry, pharmaceutical industry, petrochemical industry,pharmaceutical industry and mining industry.

A product, by-product or waste product can comprise for example whey,curd, spent grain, yeast, fungus, bacteria, mature or immature corn,potato or parts thereof, or vegetables. According to one aspect of theinvention, the product to be fermented is a product that failed to meetone or more production specifications.

Feed products according to the invention can comprise grain (ripe orunripe), such as barley, wheat, rye, oat, maize, rice, beans, peas,sorghum, triticale and/or soy.

Another aspect of the invention concerns the provision of a fermentedmixed feed using the method or methods presented above. Such a fermentedmixed feed can be provided by fermenting feeding stuff, such as cereals(mature or immature), ripe or unripe barley, wheat, rye, oat, maize,rice, beans, peas, sorghum, triticale and/or soy. Commonly, the feed tobe fermented comprises a mixture of several crops. This feed is alsodifferent according to the animals to be fed, as well as their age.Furthermore, economic considerations are also taken into account. Commoncereals lack sufficient levels of essential amino acids, such asmethionine and lysine. Soy is richer in these essential amino acids, butit is also more expensive. According to one embodiment of the inventionthe feed to be fermented comprises a mixture of barley and wheat. Inanother embodiment, the mixture comprises soy, barley, and wheat. In afurther embodiment the feed comprises maize and soy. In yet anotherembodiment, the mixture comprises barley and peas.

Another aspect of the invention relates to the use of a liquid fermentedfeed, provided as described above, for feeding animals. According to oneembodiment of the invention, the animal is a mono-gastric animal.According to a further embodiment, the animal is a pig.

A further aspect of the invention relates to the use of a liquidfermented feed as growth promoter. According to one aspect of theinvention, the animal is a pig.

In another aspect, the present invention relates to the use of afermented product as described herein directly as liquid feed or inmixture with other components such as feed products and fluids toprepare a liquid feed.

In still another aspect, the present invention relates to a method forpreparing a fermented product comprising

(a) providing a fluid product,(b) providing lactic acid-producing bacteria capable of producingsufficient lactic acid to decrease the pH to at least 4.2 within 24hours,(c) bringing (a) and (b) together, and(d) allowing fermentation to take place under suitable conditions.

In one embodiment of such method, the pH is between 4.2 and 3.5. Inanother embodiment, the pH is 3.8.

In one embodiment of the present method, the fluid product is obtainedfrom the dairy industry, the agricultural industry, the wine industry,the spirit industry, or beer industry, or combinations thereof. In oneembodiment, the fluid product is whey, maize, CCM (cob maize silage) andunriped corn, and combinations thereof.

In one embodiment of the method of the present invention, the lacticacid-producing bacteria is selected from Lactobacillus spp., Pediococcusspp., and Lactococci spp. and combinations thereof.

In a special embodiment of the method of the invention, the methodfurther comprises the addition of another feed product, and fermentationis subsequently allowed to take place under suitable conditions.

In yet another aspect, the present invention relates to a method ofpreparing a liquid feed comprising

(a) providing a liquid product according to the present invention,(b) providing a feed product to be fermented, and(c) allowing fermentation to take place under suitable conditions.

In one embodiment of the method, the liquid product is present (oradded) in a concentration of 1 vol-% or more, such as for example 1-2vol-%, or 1-5 vol-%. In another embodiment, the liquid product is addedin more than 5 vol-%. In one embodiment of the method of the presentinvention, the feed product is selected from the dairy industry, theagricultural industry, the wine industry, the spirit industry, or beerindustry, or combinations thereof. Examples are corn, maize, beans, soyand mixtures thereof.

It has surprisingly been found that the invention results in a number ofadvantages, including better preservation of the feed, improved healthof the animals resulting in decreased need of antibiotic-based growthpromoters and medicaments, improved palatability of the liquid feed, thepossibility of using other types of liquid feed, cheaper feed and apositive influence on the animal's environment.

In accordance with the present invention, the fermented product may befed directly to the animals (with or without further prior processing),or it may be used for further fermentation of liquid feed. In the lattercase, the fermented product should constitute 1 vol-% or more relativeto the total amount of feed. In another embodiment, the fermentedproduct constitutes 5 vol-%.

The present invention is based on the recognition that the lacticacid-producing bacteria should be able to produce lactic acid fastenough and in sufficient amount to out compete the natural fermentationwhich will take place due to natural occurrence of bacteria includingpathogenic bacteria and lactic acid-producing bacteria, yeasts andmoulds. It is recognised that, although many pathogenic bacteria are notable to grow at a pH lower than 4, this does not happen if theproduction of lactic acid by the lactic acid-producing bacteria takesplace quickly and efficiently. Furthermore, most yeasts and moulds areindeed able to grow under very acetic conditions, however, it has beenrecognised that quick and efficient production of lactic acid by thelactic acid-producing bacteria cause very reduced or even eliminatedgrowth of such yeasts and moulds.

It is believed that the fermented product according to the presentinvention may be prepared from any type of fluid product since thefermentation is strongly influenced by the choice of lacticacid-producing bacteria. Likewise, the fermented product of theinvention may be used for the further fermentation of any other type offeed product. Due to the advantageous properties of the fermentedproduct of the invention, the further fermentation will result in awell-controlled further fermentation, leading to a safe and tastyproduct, which furthermore has a higher nutritive value and highervitamin content. Furthermore, feeding the animals with products of thepresent invention may reduce the risk of infection with Campylobacterand Salmonella spp. and thus the present invention adds to the humanfood safety.

When water or another fluid like e.g. whey or skim milk is added duringthe fermentation process, it may advantageously be treated so as toremove or at least reduce components of the water which have buffercapacity. In particular, the content of calcium, zinc, manganese and/oriron may be reduced or removed. This is believed, in some instances, tofacilitate the effective production of lactic acid by the lacticacid-producing bacteria due to the reduce buffer capacity.

As mentioned above, pathogenic bacteria are a natural inhabitant of theanimal's surroundings, and they grow in the environment, and add to theso-called “bacterial pressure”, or “microbial pressure”. Bothexpressions are used interchangeably, and include any microorganisms,such as bacteria, yeast, fungi, amoebae, spores, phages, single-cellorganisms and the like. In accordance with the present invention, it hassurprisingly been observed that feeding the animals with the fermentedproduct of the present invention, the animal's environment is modifiedpositively.

Sometimes it is desired to initiate the fermentation under sterileconditions. This may e.g. be the case when the bacterial pressure of theenvironment is high.

However, usually lactic acid-producing bacteria, growth medium and fluidare provided together as a pre-mixed solution. A serious drawback ofthis system is that fermentation is already taking place, and thus thebacteria of the surroundings may outcompete the lactic acid-producingbacteria of the pre-mixed solution almost immediately. Furthermore, suchpre-mixed solution is difficult to ship to the customer, and it must beused within a limited period of time after it has been shipped,otherwise the bacteria cultures will die when the growth medium is usedup.

By the present invention, the problems of initiating sterile initiationof fermentation have been solved.

Thus, in an additional aspect, the present invention relates to asterile closed package system comprising a container with sterile,inactive lactic acid-producing bacteria, a container with a sterilegrowth medium for the lactic acid-producing bacteria, and a containerwith a sterile fluid. In one embodiment of the package system, thelactic acid-producing bacteria have been freeze-dried. In anotherembodiment, the growth medium comprises vitamins, maltodextrine, anddextrose anhydrose. In another embodiment, further containers may beincluded.

In another additional aspect, the present invention relates to a methodof initiating homofermentation by desired lactic acid-producing bacteriacomprising

(a) providing a sterile closed package system comprising a containerwith sterile, inactive lactic acid-producing bacteria, a container witha sterile growth medium for the lactic acid-producing bacteria, and acontainer with a sterile fluid in a container,(b) bringing the bacteria, the growth medium and the fluid within thesterile closed package system in contact with each other wherebyfermentation is initiated, and(c) bringing the content of said sterile package system into contactwith the feed product to be fermented.

In one embodiment of the method, the lactic acid-producing bacteria havebeen freeze-dried.

In yet another additional aspect, the present invention relates to theuse of a sterile closed package system as described herein forinitiating homofermentation.

The closed packaged system of the invention can be shipped to the siteof use. At the site, the individual containers are opened (e.g. bytearing, picking a hole, dissolving one or more of the containers,opening valves, melting, or pressing) allowing the components to bebrought into contact with each other under sterile conditions.

The advantages of the above method are that a homo-fermentation processcould be initiated without concerns about bacterial interference.

The containers may e.g. be made of a polymeric material e.g. plasticsand metal. Subsequently, the containers could be placed side by side inclosed package, and the package sealed.

FIG. 6 is a flow diagram, showing the steps of providing animal feedcomprising a fermented product or by-product from an industrial processaccording to an embodiment of the invention. Often, a by-product, awaste product, or a product not meeting specifications is produced in orduring an industrial produces (1). This product, here termed by-product,can be converted (2) to a fermented product, also called liquidfermented feed, by addition of lactic acid bacteria. If required, sugar,carbohydrates or a carbohydrate containing composition can be added inorder to promote growth of lactic acid bacteria. Furthermore, water canbe added in order to provide an appropriate water content. If needed,additional factors providing suitable fermentation conditions, such assalts, buffers and the like, can be added as well. Fermentation (2) cancomprise mixing, stirring and temperature control. Upon fermentation(2), a liquid fermented feed product is obtained. This liquid fermentedfeed product can be provided to animals as feed (3), either alone, or assupplement together with conventional feed, or supplemented withconventional feed.

In another embodiment of the invention, the product, by-product or wasteproduct to be fermented comprises whey, spent-grain, potatoes (or partsof potatoes (e.g. potato peel)), yeast, bacteria, or fungi, either aloneor in any combination. If appropriate, the product to be fermentedaccording to the invention can be modified chemically and/or physically,comprising e.g. treatments with heat, cold, pH, addition of chemicalcompounds and or compositions, either alone or in any combination. Thecomposition of the product to be fermented may be altered by addition ofa further product, e.g. improving nutrition value, fermentability and/orstorability of the fermented product. In a further embodiment of theinvention, a product which is not a by-product or waste product of anindustrial process is converted into a fermented product. Industrialprocesses according to the invention comprise ethanol production (e.g.beer, wine, bioethanol, distilled spirit and the like), pharmaceuticalindustry (production of pharmaceutical composition), chemical industry,agricultural and food industry (e.g. dairy, fishing, husbandry, meatprocessing) and the like.

The flow diagram shown in FIG. 7 illustrates the steps of providing afermented, liquid animal feed according to an embodiment of theinvention. Similar to Example 4, FIG. 6, a liquid fermented feed productis obtained. However, instead of feeding the liquid fermented feedproduct directly to one or more animals as in FIG. 6, the liquidfermented feed product is used to ferment (4) feed to provide afermented feed product (“fermented mixed feed”). This fermented feedproduct is the provided (5) to animals as feed, either alone, or assupplement together with conventional feed, or supplemented withconventional feed.

FIG. 8 illustrates main components and steps according to an embodimentof the invention. A indicates one or more containers comprising feed.The container(s) can comprise different feed, feed components or feedmixtures Furthermore the container(s) can comprise water or a waterbased composition. B indicates a container or fermenter comprising thefermented product. C illustrates a fermenter, wherein feed from A isprovided, mixed with fermented product from B. D illustrates either thesame fermenter as C, after fermentation, or a separate fermentater, inwhich at least a part of the fermentation of feed occurs. The arrows 1and 2 indicate providing fermenter C with feed from A and fermentedproduct from B, respectively. Arrow 3 indicates that fermention ofproduct B occurs, and/or transfer of the content or part of the contentof C into another fermenter or container. Arrow 4 indicates providingfermented feed D to animals. Means for mixing the products can beprovided in A, B, C and/or D.

In one embodiment of the invention, the animals are uniquely orpredominantly provided with fermented feed D.

In another embodiment, the animals are provided with a mixture of D andanother feed, feed component of feed mixture from A. This can bedesired, for example for compensating for negative impact offermentation on some feed ingredients, such as amino acids. It is knownthat the level of amino acids can be reduced through fermentation, whichmay not be desirable.

FIG. 9 shows a “continuous” method of producing fermented feed. “A”indicates one or more containers comprising feed. The container(s) cancomprise different feed, feed components or feed mixtures. Fermenter Bis provided with feed from container(s) A, and fermentation occurs. Thisfermentation can either occur spontaneously, driven by themicroorganisms present in the feed product to be fermented, or byproviding a starter culture in order to control the fermentation. Cindicates either B during or after fermentation, or another fermenter,in which at least a part of the fermentation takes place. The arrow atthe bottom of C indicates providing the fermented feed product toanimals. D indicates the fermenter C after a portion of the fermentedfeed product has been removed, commonly as a result of feeding animals.E indicates that the fermenter D is refilled with feed from A, and Findicates either E during or after fermentation, or another fermenter,in which at least a part of the fermentation takes place. The arrow atthe bottom of F indicates providing a portion of the fermented feedproduct to animals. The cycles D, E F are repeated. These repetitions,or “continuous” fermentation is not desirable, as the quality of thefermented feed product is known to change with time, as the compositionof the starter culture in step D will gradually change. This is forexample due to the microbial pressure, i.e. the bacteria, yeast andfungi including spores, present in or near the stable, animals, workers,feed kitchen, containers, pipes, machines, water and the like, themicrobial flora providing fermentation can change with time.Furthermore, yeast, e.g. acid tolerant yeast, is known to be a problemin “continuous” type fermentations. Although yeast is growing slowly,the levels of yeast may rise from batch to batch. Generally, yeast isundesirable, as it can provide off flavours, which e.g. pigs do notlike. Yeast infection is also a very common problem in feeding pipes,which are very difficult, if not virtually impossible to clean. Afurther potential problem associated with “continuous”-type fermentationis the risk of infections with bacteriophages. This problem is knownfrom dairy industry. Another potential issue is the occurrence ofnatural mutations, which also can reduce the quality of the fermentedproduct.

The invention, including its benefits, is further illustrated by thefollowing non-limiting examples.

EXAMPLES Example 1 Preparation of a Fermented Product According to thePresent Invention

The present invention was exemplified by mixing 80% wheat and 20% barleyin a fermentation tank. In the mixture, grain constituted 28% and waterthe remaining 72%. Furthermore, 100 parts of Pig Stabilizer 317(Medipharm) were added per 10 tones of liquid feed soup. The PigStabilizer 317 further comprised vitamins, maltodextrine, dextroseanhydrose and a combination of the lactic acid-producing bacteriaPediococcus pentosaceus, Pediococcus acidilactici and Lactobacillusplantarum. The air intake to the fermentation tank was taken directlyfrom the air in the room wherein the fermentation tank was situated.Samples were sent to a commercial analytical laboratory, and analysisfor vinegar and ethanol content indicated that the environment wasalmost sterile since the analysis only revealed 3 mM of vinegar and noethanol in the mixture. Furthermore, the level of lactic acid was about140 mM after 20 hours fermentation. This mixture could thus beconsidered almost as 100% homofermentative.

In conclusion, it is possible to prepare a fermented product usinglactic acid-producing bacteria. Furthermore, it has been demonstratedthat fermentation is controlled and takes place in a homofermentativeway.

Example 2 Preparation of Feed Products According to the PresentInvention

Samples of the fermented product of Example 1 were taken and brought toa liquid feed kitchen where liquid feed had previously failed to beproduced due to a high bacterial pressure of the surroundings.

Experiment 1

1 liter of the mixture was mixed with 100 liters of feed product in thefermentation tank in the kitchen. The fermentation process was completedafter 9 hours and the pH of the mixture was 3.8.

Experiment 2

In another example, 50 liter of the mixture was mixed with 200 liters offeed product in the fermentation tank in the kitchen. The fermentationprocess was completed after 5 hours and the pH of the mixture was 3.8.

Experiment 3

In third example 50 liter of the mixture was mixed with 5000 liters offeed product in the fermentation tank in the kitchen. The fermentationprocess was completed after 9 hours and the pH of the mixture was 3.8.

Results

In all instances, the obtained liquid feed was almost homofermented.Thus, it has been demonstrated that the fermentation process of thepresent invention has superior advantages since in had previously provedimpossible to obtain a homofermented liquid feed. It is indeed possibleto control the fermentation even when the bacterial pressure of thesurroundings is extremely high.

Example 3 Preparation of Liquid Feed in Feed Kitchens Having LowerBacterial Pressure

Liquid feeds similar to ones described in Example 2 were prepared inother feed kitchens with a lower bacterial pressure of the surroundings.The fermentation time as compared to the fermentation time in Example 2varied within 1 hour.

Accordingly, it was demonstrated that the fermentation according to thepresent invention takes place so quickly and so efficiently that thebacterial pressure does not influence the outcome of the fermentationsubstantially.

Example 4 Different Set-Ups of Home Mixer Feed Systems (Systems 1-3)

FIGS. 10-12 illustrated 3 different home mixer feed systems, suitablefor providing a fermented mixed feed product. Routinely, fermentation ofthe feed product to be fermented will be finished after about 6 h atbetween 20-25° C., preferably around 23° C. A home mixer feed system cancomprise several of the following components:

A) Fermentation tank A which will weigh ground mixture of any fermentedproduct, feed and water from C, F and E. Commonly, there will be takenfrom 5-70% fermented product from F. After about 6 hours at ±23° C., thefermentation will be finished. However, temperatures ranging from 15° C.to 45° C. can also provide fermentation). Commonly, the size of thefermentation tank A is from 1.000 kg, upwards.B) Mixing feeding and fermentation tank B, mixing content from A (ifinstalled), C, D and E. Commonly, the size of mixing tank B is rangingfrom 500 kg to 12000 kg. Fermented mixed feed is best used fresh, orwithin 24 h. If fermented grain is standing too long after fermentation(e.g. two or more days), the fermented feed product can turn bitter,presumably due to the high level of minerals in the grain and otherchemical reactions.C) One or more tanks C containing feed product to be fermented.Commonly, they contain grain, such as wheat and barley, as well as soy.Often, the feed product to be fermented is not heat treated and notsterile.D) Vitamins and minerals containers D. These components can be providedas one or more mixtures, or individually. They can be provided in bulkor big bags. Mostly, dry components (powder, pellets and the like) areused, but they can also be liquid. Commonly, amino acids are added afterthe fermentation process, in order to avoid degradation, decompositionand/or destruction of the amino acids.E) Clean soft water tank E, providing water at 20-25° C., preferablyaround 23° C. The water temperature can also be different from roomtemperature, e.g. lower than or at around ground temperature. This isnot uncommon, if water from tank E is added into fermentation tank B.Water is checked for minerals, and if necessary, adjusted and/or cleanedto obtain the best growth for the lactic acid bacteria. The water tank Ecan have any capacity, ranging from very small to very large.Alternatively, water is provided directly from a tap. A heat-exchangercan be installed to heat up the water if necessary.F) Tank for fermented product with lactic acid bacteria. Often, thecapacity of fermented product tank F is from 1000 kg, upwards. Normally,the fermented product remains active and usable for a storage time of upto 2 weeks. Commonly, the tank is emptied and cleaned before fillingwith a new batch of fermented product. The tank can be provided with azip system to facilitate emptying and cleaning. Alternatively, the tankis emptied and cleaned by conventional means.G) Feeding pipes G bringing the feed to the different sections in thepig house. Feeding pipe G can be several 100 m long, and they can reachlength of 1 km or more. Depending on the age of pigs they need differentenergy and mineral levels in the feed. Therefore there can be severallines of different pipes, e.g. one for each section, or one main pipewhere the feed in the pipe is mixed with the new blended feed, so allthe pigs get the same type of feed. It can also be a pipe, where thefeed is emptied with water. Feeding pipe G can be a return pipe, where afraction of the feed is returned to the fermenting tank B. It can alsobe a one-way pipe. Different sections of the stables can have individualfeeding pipes.

System 1

FIG. 10 shows an embodiment of the current invention with a liquidfeeding system comprising a fermentation tank A and afermentation/mixing tank B. Feeding pipe G can return the liquidfermented mixed feed into fermenter B.

System 2

FIG. 11 shows another embodiment of the current invention with a liquidfeeding system comprising a fermentation/mixing tank B, but withoutfermentation tank A. Feeding pipe G can return the liquid fermentedmixed feed into fermenter B; alternatively, the mixed feed istransported only one way. All feed components are mixed in tank B.Generally, amino acids from tank D are added towards the end offermentation, preferably before feeding. Also water (preferably 23° C.,but also cold groundwater) can be added to fermenter B.

System 3

FIG. 12 shows an alternative embodiment of the current invention. Theliquid feeding system comprises several fermentation tanks A, wherewater from E, the fermented liquid fermented food from F and the feedproduct to be fermented (C) can be provided individually to severalparallel fermenters A.

Example 5

Feeding Trials with Different Ratios of Liquid Fermented Product: FeedProduct to be Fermented

The results of feeding trials using different liquid fermented productsand different home mixer feed systems are presented below. The resultsare based on experiments conducted at 19 different pig farms in Europe.

Fifteen farmers used fermented potatoes as inoculum for fermentation offeed products for pig production. Fourteen sites were in Jutland,Denmark, and 1 site was located in Wasa, Finland. Three sites had a homemixer feed systems 1 (FIG. 10), 11 sites had a home mixer feed system 2(FIG. 11), and 1 site had a home mixer feed system 3 (FIG. 12).

The feeding trials were performed on sows, weaners and finishers, andtypically, the pigs weight ranged from ˜7 to ˜200 kg.

Generally, the farms were comparatively large, and had more than 250animal units per farm or more (1 animal unit equals approximately 32finisher pigs per year from 30 to 107 kg live weight).

On three farms, fermented whey was tested. The sites were in thesouthern part of UK; Wasa, Finland; and Jutland, Denmark, respectively.All three farms possessed a home mixer feed system 2 withoutfermentation tank A (FIG. 11).

On one site on Zealand, Denmark (home mixer feed system 1, FIG. 10),feeding pigs with fermented spent grain from beer production andfermented molasses was tested. This trial was only conducted withfinishers.

The liquid fermented product used was fresh potato peals from a potatofactory. Commercially available lactic acid bacteria starter cultures,either “PIG stabiliser 317” (Medipharm, Sweden), or “Pig Stabiliser 400”(Lactosan, Austria) were added in the factory after the pealing machine,where the temperature was about 38° C. Approximately 3 g lactic acidbacteria were added per 1000 kg of potato peals. The dry matter contentwas about 12.5 to 16%. The potato peals were pumped to astorage/fermentation tank made of fibreglass, with a capacity of around50 m³ without stirring. In the factory, where the lactic acid bacteriawere sprayed onto the potato peals, the temperature was 38° C. When thepeals arrived into the storage tank the temperature dropped to about 35°C. and kept for 24 hours. After fermentation in the storage tank,solanine from the potatoes destroyed. Furthermore, the digestibility ofthe potato peals was increased compared to non-fermented potato peals(data not shown). The fermented potato peals (=liquid fermented product)was transported to the locations in a tank lorry, and transferred/pumpedinto tank F at the different test sites.

The fermented mixed feed was freshly fermented and used within 24 hoursafter fermentation. Generally, it consisted of grains and soy. Allfeeding rations comprised mainly wheat, barley and soy. The maximum soycontent was 23% of total. In 14 rations wheat was the dominantingredients used from 40% to 75% of total. In 1 ration, barley was thedominant ingredient.

In the fermentation tank (tank A) the dry matter content was from 29 to32% as the distance to pump it to tank B was always less than 10 metres.The dry matter content in tank B was from 24% to 27% depending on thedistance to pump the feed inside the pig house.

Generally, feeding trials with fermented mixed feed were performed onthe whole pig stock. The results presented below are based on acomparison of performance data prior to installation of the home mixerfeed systems, and after installation and feeding with fermented mixedfeed.

Results:

The feeding trial results are summarized in Table I below:

TABLE I Results from feeding trials Mixture Condition 1 Condition2 %inoculum low microbial pressure high microbial pressure (liquid (no pigshad been kept in (pigs had been kept in stables fermented stables priorto feeding prior to feeding experiment) product) experiment)  5% 2trials were perfect(*) 1 trial was perfect 1 trial negative(**) 2 trialswere negative 10% 3 trials were perfect 2 trials were perfect 2 trailswere negative 30% 3 trials were perfect 3 trials were perfect 1 trailwas negative 40% 3 trials were perfect 3 trials were perfect(*)“Perfect” indicates that pigs were eating very rapidly, and they wereeating essentially all the feed provided to them, and quicker thanbefore (i.e. conventional feeding without provision of liquid fermentedmixed feed). Daily weight gain was either as before or increased, e.g.at 0-50 g per day. If 900 gram daily gain was achieved before startingthe feeding trials, daily weight gain increased from 900-950 g per day.No or little diarrhoea was observed, requiring no medicine/antibiotics.Furthermore, the feed conversion ratio (FCR) was reduced byapproximately 0.2 compared to before the feeding trial. FCR, also calledfeed conversion efficiency (FCE), is a measure of an animal's efficiencyin converting feed mass into increased body mass. Specifically FCR isthe mass of the food eaten divided by the body mass gain, all over aspecified period of time. FCR is dimensionless. Animals that have a lowFCR are considered efficient users of feed. (**)“Negative” indicatesthat the pigs did not want to eat the estimated amount of feed given tothem. By visual inspection it could be seen that pigs suffered fromdiarrhoea (more than 20%), and that the pigs did not grow well(estimated 20 to 60 g less weight increase per day). If 900 gram dailyweight gain was achieved before feeding with fermented mixed feed, dailyweight gain dropped to 840-880 gram daily. As a result, antibiotics hadto be used for treating stomach problems. Thereby acceptable weightgains could be re-established.

It is clear that type and microbial pressure in the surroundings (house,farm, etc) where the fermentation is done, has a great influence on theamount of liquid fermented product needed to be added to the feedproduct to be fermented, in order to provide a successful and reliablefermentation. Higher microbial pressures require higher inoculum ofliquid fermented product.

It is a known problem that the feeding line G cannot be cleanedadequately. The feeding line comprises e.g. a lot of rubber valves,which are difficult to access and which cannot be cleaned, and whichoften results in infection and growth of yeast. This risk is increasedin feeding lines, where feed from the feeding pipe G is returned in tothe fermenter, which may comprise acid tolerant (and undesired) yeast.However, when using 30% or more of liquid fermented product asincoculum, this problem can be avoided, as feeding with fermented mixedproduct according to the invention proved successful. Generally, in newhouses/stables there was no built up of yeast was observed and feedingtrials were successful.

Smaller pigs were more sensitive against taste of the feed (data notshown).

The fermentation tank has little influence on the fermentation. Nodifferences were observed between the different liquid feeding systemsor brands thereof.

Fermenter systems supplied by Funky (Hammerum near Herning, Denmark),Skiold (Ikast, Denmark) and Big Dutchman (Vejen, Denmark) were used withsuccess. Funky and Big Dutchman tanks are made from glass fibre. Two ofthe tanks from Skiold are made from stainless steel, and one from glassfibre. All the feeding tanks (B) were made from glass fibre and thesizes varied from 5 to 8000 litres.

Likewise, rotation speeds of the mixing means used during fermentationwere found not to be critical. The mixing/stirring systems used werethose provided with the fermentation tanks. Different rotation speed,ranging from about 35 rpm to about 250 rpm were used, without observingdifferences in the final results.

The startercultures “PIG stabiliser 317” and “Pig Stabiliser 400”comprised rapidly acidifying, homofermentative strains of lactic acidbacteria (Pediococcus pentosaceus HTS (LMG P-22549), Pendiococcusacidilactici NCIMB 30086 and Lactobacillus plantarum LSI (NCIMB 30083);Enterococcus fæcium NCIMB 30122, Lactobacillus rhamnosus NCIMB 30121)that were able to produce a lot of lactic acid within a short time. Ithas proven earlier that the inoculum can produce amounts of lactic acidexceeding 160 mM. It is also important to use a strain with goodpalatability for the pigs. In general, it was important that the rightstrain of lactic acid was used, to obtain a rapid drop in pH under pH4.0. Thereby, a preservation of the feed is obtained and destruction ofamino acids is reduced. No differences were noticeable between the twostarter cultures used.

The fermented feed product and its composition were found not to be ofgreat importance. There were no differences in the observed results,regardless if barley or wheat was the dominant ingredients. However, theamount of soy in the feed product had an influence on pH and lacticacid. A higher amount of Soy used as more lactic acid. Interestingly, anincrease of 15% in digestibility of soy was observed, when feeding withfermented mixed feed comprising soy.

In conclusion, feeding pigs with a fermented mixed feed provides thefollowing benefits (this list is not to be construed as to be limitingfor the invention):

-   -   higher feed intakes for the pigs    -   better palatability of the feed    -   higher growth rates of the pigs    -   improved nutritial value of the feed—the nutritial value of the        fermented mixed feed is higher than the nutritional values of        the liquid fermented product and the feed product to be        fermented (without fermentation).    -   control/preservation of amino acids, avoiding degradation of the        free amino acids. This was only obtained were 10% or more        fermented products was added into the feed.    -   digestibility of soy protein was increased with about 15%.    -   use as growth promoter—improved or perfect stomach health        reduced or removed the need for medicine against E. coli,        Salmonella and/or Lawsonia diarrhea.    -   improved stomach health through fermented mixed feed—this allows        for selecting and using grain/feed which is cheaper.    -   robust pigs—pigs are less sensitive towards change in feed    -   financial benefits—reduced medical expenses, reduced expenses        for feed, increased weight gain of the pigs.    -   robust and reliable liquid feeding system—feeding animals with        fermented mixed feed is now feasible, where previous systems        sometimes or often failed. The system is farmer friendly, and        manageable in an environment with high microbial and/or        bacterial pressure. Undesired growth of yeast in e.g. the        feeding pipe G can be avoided. In contrast to natural        fermentation systems, the microbial flora can be controlled.        Furthermore, the system does not require sterile or near-sterile        conditions, adding to its reliability and robustness. Due to the        use of the liquid fermented product as inoculum, which comprises        a high number of active bacteria, active bacteria are added to        the feed product to be fermented. This reduces the lag-phase of        the fermentation, resulting in a rapid production of lactic acid        and reduction of pH.

Example 6 Preparation of a Mixed Fermented CCM Silage

A mixed fermented product produced according to Example 5, obtained byfermenting a mixture of 50% wheat, 40% barley with 10% soy, was pumpedinto 12 1000 liter pallets containers made of plastic. The palletcontainers were stored at ambient temperature without any stirring forup to 1 week. Over a period of 2 weeks, newly combined CCM maize wasinoculated with the mixed fermented product by pumping and spraying themixed fermented product onto the CCM maize, while it was transported viaan auger into the storage silo. The storage silo was a conventionalupright standing 1200 t airtight silo, presumably made of coated steel(brand unknown). The auger was about 15 m long with a diameter of around200 mm. The ratio of mixed fermented product: CCM maize wasapproximately 1-2%. The silo was filled within 14 day with CCM maizeinoculated with the fermented mixed product and closed.

After approximately 6 month, the silo was opened and the now fermentedCCM was emptied using an auger. A sample of ˜0.5 l was removed, and thesample was send to an analytical laboratory. Analysis revealed lowlevels of acetic acid (10.5 mM), and a lactic acid concentration of 110mM.

The fermented CCM was used in one of the feeding trials described inExample 5, and was labelled “perfect”. It is a known problem that pigsdo not like acetic acid. However, the levels of lactic acid were so lowthat it was possible to feed pigs (˜30 kg body weight) with thefermented CCM, as they were not reluctant to eat the fermented CCM.

Conventional CCM-based silage are known to have varying levels of aceticacids, commonly ranging between for example 20-200 mM acetic acid ormore. These variations are assumed to be caused by the water content ofthe CCM maize during harvest and the microbial flora present duringfermentation.

The example presented above demonstrates that a mixed fermented productbased on fermented CCM maize can be produced, and the CCM maize ensilageis suitable for feeding monogastric animals such as pigs.

1. A method of preparing a fermented mixed feed, wherein said methodcomprises the steps of: (a) providing a liquid fermented product byfermentation with lactic acid bacteria; (b) providing a feed product tobe fermented; (c) combining the products from steps (a) and (b), andfermenting the feed product of step (b) using the liquid fermentedproduct of step (a) as inoculum; wherein the inoculum comprises a highnumber of active lactic acid bacteria providing a rapid production oflactic acid and reduction of pH in step (c).
 2. A method according toclaim 1, wherein the fermentation in step (a) is essentially ahomofermentative fermentation.
 3. A method according to claim 1, whereinwherein the fermentation in step (c) is essentially a homofermentativefermentation.
 4. A method according to claim 1, wherein the method ofpreparing a liquid fermented feed is suitable for an environment of highmicrobial pressure.
 5. A method according to claim 1, wherein afermented mixed feed is provided with a pH of 4.2 or lower.
 6. A methodaccording to claim 1, wherein and said fermented mixed feed has a lacticacid concentration in the range of 50-300 mM.
 7. A method according toclaim 1, wherein said fermented mixed feed is prepared within a timeperiod of up to 24 hours.
 8. A method according to claim 1, wherein thefermentation step (c) is conducted at a temperature between 10-50° C. 9.A method according to claim 1, wherein mixing means are provided duringthe fermentation step (c), said mixing means operating at between 1-500rpm.
 10. A method according to claim 1, wherein in the combined productprovided in step (c), the proportion of liquid fermented product is inthe range of 0.1 to 99.9 vol-%.
 11. A method according to claim 1,wherein the pH of the liquid fermented product provided in step (a) isless than 4.2.
 12. A method according to claim 11, wherein the lacticacid-producing bacteria comprise lactic acid bacteria selected from thegroup consisting of one or more of Enterococcus spp., Lactobacillusspp., Lactococcus spp., and Pediococcus spp.
 13. A method according toclaim 1, wherein the liquid fermented product provided in (a) has beenobtained by fermentation with an inoculum comprising lactic acidbacteria selected from the group consisting of one or more ofEnterococcus spp., Lactobacillus spp., Lactococcus spp., and Pediococcusspp.
 14. A method according to claim 12, wherein the lactic acidbacteria are selected from the group consisting of one or more ofEnterococcus faecium, Lactobacillus rhamnosus, Lactobacillus plantarum,Pediococcus acidililactili, and Pediococcus pentosaceus.
 15. A methodaccording to claim 1, wherein the liquid fermented product provided in(a) is obtained by fermenting one or more industrial products.
 16. Amethod according to claim 15, wherein the industrial product is aby-product or a waste product.
 17. A method according to claim 16,wherein the by-product or the waste product is one or more of whey,curd, spent grain, yeast, fungus, bacteria, plants or parts thereof,potato or parts thereof.
 18. A method according to claim 1, wherein theliquid fermented product provided in step (a) is provided by fermentinga product comprising one or more of whey, curd, spent grain, yeast,fungus, bacteria, mature or immature corn, potato or parts thereof, orvegetables.
 19. A method according to claim 1, wherein the feed productprovided in step (b) comprises one or more of ripe or unripe plants orparts thereof.
 20. A method according to claim 1, wherein the feedproduct provided in step (b) comprises one or more of barley, wheat,rye, oat, maize, rice, beans, peas, sorghum, triticale and soy.
 21. Amethod according to claim 1, wherein the fermented mixed feed is silage.22. A method according to claim 21, wherein said silage is preparedwithin up to 1 day.
 23. A method according to claim 21, wherein theproportion of liquid fermented product provided in step (c) is in therange of 1-2 vol-%.
 24. A method according to claim 21, wherein the feedproduct to be fermented is CCM maize.
 25. A silage prepared by a methodaccording to claim
 21. 26. A fermented mixed feed prepared by a methodaccording to claim
 1. 27. A fermented mixed feed according to claim 26,wherein the feed provided in step (b) comprises one or more of ripe orunripe plants or parts thereof.
 28. A fermented mixed feed according toclaim 27, wherein the feed provided in step (b) comprises one or more ofbarley, wheat, rye, oat, maize, rice, beans, peas, sorghum, triticaleand soy.
 29. A fermented mixed feed according to claim 26, wherein thefeed provided in step (b) comprises a mixture of: barley and wheat; soy,barley, and wheat; maize and soy; or barley and peas.
 30. A method forfeeding an animal which comprises administering to the animal afermented mixed feed prepared by: (a) providing a liquid fermentedproduct by fermentation with lactic acid bacteria; (b) providing a feedproduct to be fermented: (c) combining the products from steps (a) and(b), and fermenting the feed product of step (b) using the liquidfermented product of step (a) as inoculum;
 31. A method of feeding ananimal according to claim 30, wherein the animal is a monogastricanimal.
 32. A method of promoting growth in an animal which comprisesadministering to the animal a feed prepared according to claim 1 as agrowth promotor.
 33. The method of claim 31, wherein the animal is apig.
 34. A method of feeding a pig which comprises administering to thepig a silage prepared according to claim
 25. 35. A method according toclaim 5, wherein a fermented mixed feed has a pH of less than 4.2
 36. Amethod according to claim 5, wherein a fermented mixed feed has a pHbetween 4.2 and 3.5.
 37. A method according to claim 5, wherein afermented mixed feed has a pH of about 3.5.
 38. A method according toclaim 6, wherein mixing means are provided during the fermentation step(c), said mixing means operating at 1-500 rpm, 10-300 rpm, or 35 1 100rpm said fermented mixed feed has a lactic acid concentration in therange of 50-250 rpm.
 39. A method according to claim 6, wherein mixingmeans are provided during the fermentation step (c), said mixing meansoperating at 1-500 rpm and said fermented mixed feed has a lactic acidconcentration in the range of 100-150 nM.
 40. A method according toclaim 6 wherein mixing means are provided during the fermentation step(c), said mixing means operating at 1-500 rpm and said fermented mixedfeed has a lactic acid concentration in the range of 250-300 nM.
 41. Amethod according to claim 7 said fermented mixed feed is prepared withina time period of up to 4-24 hours.
 42. A method according to claim 7said fermented mixed feed is prepared within a time period of up to12-24 hours.
 43. A method according to claim 7 said fermented mixed feedis prepared within a time period of less than 4 hours.
 44. A methodaccording to claim 8 wherein the fermentation step (c) is conducted at atemperature between 15-40° C.
 45. A method according to claim 8 whereinthe fermentation step (c) is conducted at a temperature between 18-30°C.
 46. A method according to claim 8 wherein the fermentation step (c)is conducted at a temperature between 22-24° C.
 47. A method accordingto claim 9 wherein mixing means are provided during the fermentationstep (c), said mixing means operating at between 10-300 rpm.
 48. Amethod according to claim 9 wherein mixing means are provided during thefermentation step (c), said mixing means operating at between 35-100rpm.
 49. A method according to claim 10 wherein in the combined productprovided in step (c), the proportion of liquid fermented product is inthe range of 1 to 99 vol-%.
 50. A method according to claim 10 whereinin the combined product provided in step (c), the proportion of liquidfermented product is in the range of 5-70 vol-%.
 51. A method accordingto claim 10 wherein in the combined product provided in step (c), theproportion of liquid fermented product is in the range of 10-50 vol-%.52. A method according to claim 11 wherein the pH of the liquidfermented product provided in step (a) is about 3.8.
 53. A methodaccording to claim 11 wherein the pH of the liquid fermented productprovided in step (a) is between 3.5 and 4.2.
 54. A method according toclaim 22, wherein said silage is prepared within up to 1 week.
 55. Amethod according to claim 22, wherein said silage is prepared within upto 1 month.
 56. A method according to claim 22, wherein said silage isprepared within up to 6 months.
 57. A method according to claim 22,wherein said silage is prepared within up to 1 year.